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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
16/03/2011 - 18/03/2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study according to international guideline (OECD guideline 202) under GLP. No deviations from guideline reported.
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Principles of method if other than guideline:
Not relevant
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not relevant
Analytical monitoring:
yes
Details on sampling:
- Concentrations: nominal loading rates (WAF) of 1.0, 1.8, 3.2, 5.6 and 10 mg/l
- Sampling method: duplicate water samples were taken after 0 h (fresh media) and 48 hours (old media)
- Sample storage conditions before analysis: stored at -20 ºC until further analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Amounts of test item (23 and 41.4 mg) were added separately to the surface of 23 litres of reconstituted water in separate stirring vessels with minimal headspace to give the loading rates of 1.0 and 1.8 mg/I. Similarly, amounts of test item (35.2, 61.6 and 110 mg) were added separately to the surface of 11 litres of reconstituted water in separate stirring vessels with minimal headspace to give the loading rates of 3.2, 5.6 and 10 mg/I. After the addition of the test item, the reconstituted water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixture allowed to stand for 1 hour. A wide bore glass tube, covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed through the Nescofilm seal. The aqueous phase or WAF was removed by mid-depth siphoning (the first approximate 75-100 ml discarded) to give the 1.0, 1.8, 3.2, 5.6 and 10 mg/I loading rate WAFs.
- Controls: Yes
- Evidence of undissolved material (e.g. precipitate, surface film, etc): Microscopic inspection of the WAFs showed no microdispersions or undissolved test item to be present.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Source: in-house labaratory strain
- Age at study initiation (mean and range, SD): <24 hours
- Method of breeding: Adult Daphnia were maintained in 150 ml glass beakers containing approximately 100 ml of Elendt M7 medium (see Appendix 1) in a temperature controlled room at 21 to 22°C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn. and dusk transition periods. Each culture was fed daily with a mixture of algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension. Culture conditions ensured that reproduction was by parthenogenesis.
- Feeding during test: no

ACCLIMATION
- Acclimation period: none
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
Not relevant
Hardness:
250 mg/l CaCO3
Test temperature:
21 - 22 ºC
pH:
7.7 - 8.0
Dissolved oxygen:
7.1 - 9.2 mg/l
Salinity:
Not relevant
Nominal and measured concentrations:
Nominal WAF loading rates: 1.0, 1.8, 3.2, 5.6 and 10 mg/l
Measured TOC corrected for control (0 hours): 1.02, 1.98, 2.82, 2.71 and 6.33 mg/l
Measured TOC corrected for control (72 hours):
Details on test conditions:
TEST SYSTEM
- Test vessel: conical flask
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: glass, 100 ml filled with 100 ml of medium
- Aeration: No
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water
An aliquot (25 ml) of each of solutions a-d was added to each litre (final volume) of deionised water with a conductivity of <5 pS cm"'. The reconstituted water had a pH of 7.8 ± 0.2 adjusted (if necessary) with NaOH or HCI and was aerated until the dissolved oxygen concentration was approximately air-saturation value.

Stock Solutions

a) CaCI2.2H20 11.76 g/l
b) MgSO4.7H20 4.93 g/l
c) NaHCO3 2.59 g/l
d) KCI 0.23 g/l

Solution Concentration of stock solution (g/l)
(XV)CaCI2.2H20 293.80
(XVI)NaHCO3 64.80
(XVII)MgSO4.7H20 246.60
(XVIII)Na2SiO3.9H20 50.00
(XIX)KCI 58.00
(XX)NaNO3 2.74
(XXI)K2HPO4 1.84
(XXII)KH2PO4 1.43

Vitamin Nutrients
Solution Concentration ofstock solution (mg/l)
(XXIII) Thiamine hydrochloride 750
Cyanocobalamine (vitamin B12) 100
D(+) biotin (vitamin H) 75

Solution Concentration of stock solution (mg/1)
(I) H3B03 57190
(II) MnC12.4H20 7210
(III) LiCl 6120
(IV) RbCI 1420
(V) SrC12.6H20 3040
(VI) NaBr 320
(VII) Na2MoO4.2H20 1260
(VIII) CuC12.2H20 335
(IX) ZnC12 260
(X) CoCI2.6H20 200
(XI) KI 65
(XII) Na2SeO3 43.8
(XIII) NH4VO3 11.5
(XIV) Na2EDTA.2H20 5000
FeSO4.7H20 1991

- Conductivity: < 5 uS/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement: after 0 and 48 hours

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light / 8 hours dark with 20 min dusk/dawn periods

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobilisation (every 24 hours)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Range finding study: yes
- Test concentrations: 1.0, 10 and 100 mg/l
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
5 mg/L
Nominal / measured:
nominal
Conc. based on:
other: Water accomodated fraction (WAF)
Basis for effect:
mobility
Remarks on result:
other: 95%CL: 4.8 - 8.4 mg/l
Duration:
24 h
Dose descriptor:
EL0
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
other: Water accomodated fraction (WAF)
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
1.6 mg/L
Nominal / measured:
nominal
Conc. based on:
other: Water accomodated fraction (WAF)
Basis for effect:
mobility
Remarks on result:
other: 95%CL: 1.4 - 1.8 mg/l
Duration:
48 h
Dose descriptor:
EL0
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
other: Water accomodated fraction (WAF)
Basis for effect:
mobility
Details on results:
- Mortality of control: 0%
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: Observations on the test media were carried out during the mixing and testing of the
WAFs. At both the start and end of the mixing period and following the 1-Hour standing period the 1.0, 1.8, 3.2 and 5.6 mgll loading rate WAFs were observed to have formed clear colourless water columns with globules of test item floating at the surface. The 10 mg/I loading rate WAF was observed to have formed a clear colourless water column with globules of test item floating at the surface and dispersed throughout at both the start and end of the mixing period and following the 1-Hour standing period. Microscopic inspection of the WAFs showed no micro-dispersions or undissolved test item to be present. At the start and throughout the duration of the test all control and test preparations were observed to be clear colourless solutions.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Mortality:
- 48h-EC50/LC50: 1.5 mg/l
Reported statistics and error estimates:
The EL*50 value and associated confidence limit at 24 hours and the slope of the response curve and standard error were calculated by the maximum-likelihood probit method (Finney 1971) using the ToxCalc computer software package (ToxCalc 1999) and at 48 hours the EL*50 value and associated confidence limits were calculated by the trimmed Spearman-Karber method (Hamilton et al 1977) using the ToxCalc computer software package (ToxCalc 1999) Probit analysis is used where two or more partial responses to exposure are shown. When only one partial response is shown the trimmed Spearman-Karber method is appropriate.

For detailed results, see attached file "Results.doc".

Validity criteria fulfilled:
yes
Remarks:
Mortality in control <10%, dissolved oxygen > 3 mg/l
Conclusions:
The acute toxicity (48h-EL50) of cinnamon leaf oil towards Daphnia magna in a WAF test is 1.6 mg/l.
Executive summary:

The acute toxicity of cinnamon leaf oil towards Daphnia magna was investigated according to OECD guideline 202 under GLP. Daphnids were exposed to nominal WAF loading rates of 1.0, 1.8, 3.2, 5.6 and 10 mg/l and observed for 48 hours. Based on nominal loading rates the 48h-NOEL and 48h-EL50 were found to be 1.0 and 1.6 mg/l respectively.

Description of key information

The result from the key study using WAFs with Cinnamon Leaf Oil was 48h-EL50 = 1.6 mg/l

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
1.6 mg/L

Additional information

The acute toxicity of cinnamon leaf oil towards Daphnia magna was investigated according to OECD guideline 202 under GLP. Daphnids were exposed to nominal WAF loading rates of 1.0, 1.8, 3.2, 5.6 and 10 mg/l and observed for 48 hours. Based on nominal loading rates the 48h-NOEL and 48h-EL50 were found to be 1.0 and 1.6 mg/l respectively.

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